Disease relevance of GUCA1A

• Recombinant GCAP was functionally expressed in Escherichia coli [1].
• METHODS: The GCAP1 and GCAP2 genes were screened by direct sequencing for mutations in 216 patients and 421 patients, respectively, with various hereditary retinal diseases [2].

High impact information on GUCA1A

• Molecular cloning and characterization of retinal photoreceptor guanylyl cyclase-activating protein [3].
• Guanylyl cyclase-activating protein (GCAP) is thought to mediate Ca(2+)-sensitive regulation of guanylyl cyclase (GC), a key event in recovery of the dark state of rod photoreceptors following light exposure [3].
• We have found that binding of Mg(2+) dramatically affects both Ca(2+)-dependent conformational changes in GCAP-1 and Ca(2+) sensitivity of RetGC regulation by GCAP-1 and GCAP-2 [4].
• No Ca(2+)-dependent oligomerization of GCAP1 was observed at physiologically relevant Ca(2+) concentrations, in contrast to the observation reported by others for GCAP2 [5].
• Here, we investigated changes in GCAP1 structure using mutagenesis, chemical modifications, and spectroscopic methods [5].

Biological context of GUCA1A

• We isolated GCAP from bovine rod outer segments, determined amino acid sequences of proteolytically obtained peptides, and cloned its gene [1].
• Guanylyl cyclase activating protein. A calcium-sensitive regulator of phototransduction [6].
• Guanylyl cyclase-activating proteins (GCAP) are EF-hand Ca(2+)-binding proteins that activate photoreceptor guanylyl cyclase (RetGC) in the absence of Ca(2+) and inhibit RetGC in a Ca(2+)-sensitive manner [4].
• This is that the GCAP2 module transmits the Ca(2+) signals to the catalytic domain from its C-terminal side and the GCAP1 module from the distant N-terminal side [7].
• The myristoyl group in GCAP-1 had a strong influence on the Ca2+-dependent regulation of ROS-GC1 (shift in IC50) [8].

Anatomical context of GUCA1A

• Activation of guanylyl cyclase in vertebrate photoreceptor cells by native acylated GCAP was half-maximal at 100 nM free [Ca2+] with a Hill coefficient of 2 [1].
• Using immunocytochemical and biochemical methods, we show here that GCAP1 is present in rod and cone photoreceptor outer segments where phototransduction occurs [6].
• We applied surface plasmon resonance (SPR) spectroscopy to monitor the Ca2+-dependent binding of myristoylated and nonmyristoylated GCAP-1 and GCAP-2 to immobilized phospholipid membranes [8].
• In this communication we show that, like the native system, ROS-GC expressed in COS cells is activated 4-6-fold by recombinant GCAP1 at 10 nM Ca2+ and that the reconstituted system is inhibited at physiological levels of Ca2+ (1 microM) [9].
• Here we report for the first time detection of a 19 kDa protein (p19) with GCAP properties in extracts of rat retina and pineal gland [10].

Associations of GUCA1A with chemical compounds

• In certain pinealocytes, ROS-GC1 coexisted with its other Ca(2+) modulator, guanylate cyclase activating protein type 1 (GCAP1) [11].
• Lowering free Mg(2+) concentrations ([Mg](f)) from 5.0 mm to 0.5 mm decreases the free Ca(2+) concentration required for half-maximal inhibition of RetGC ([Ca]((1/2))) by recombinant GCAP-1 and GCAP-2 from 1.3 and 0.2 microm to 0.16 and 0.03 microm, respectively [4].
• Two Cys residues of GCAP1 situated in spatially distinct regions of the N-terminal domain (positions 18 and 29) and two Cys residues located within the C-terminal lobe (positions 106 and 125) were employed to detect conformational changes upon Ca(2+) binding [5].
• The myristoyl group restricted the accessibility of one cysteine in GCAP-1 and GCAP-2 observed by measuring the time-dependent thiol reactivity of cysteines [8].
• We conclude that myristoylation of GCAP-1 and GCAP-2 is important for fine tuning of guanylate cyclase activity [12].

Other interactions of GUCA1A

• We propose that p24 be referred to as GCAP-2 and that GCAP be referred to as GCAP-1 [13].
• These results demonstrate that GCAP1 is an activator of ROS GC, while the finding of a second activator, GCAP2, suggests that a similar mechanism of GC regulation may be present in outer segments, other subcellular compartments of the photoreceptor, or other cell types [6].
• The results suggest that GCIP is a Ca2+-binding protein of the GCAP/recoverin subfamily [14].
• It represents a newly identified family of neuronal specific Ca(2+)-binding proteins that includes neurocalcin, hippocalcin, and guanylyl cyclase-activating protein [15].

Analytical, diagnostic and therapeutic context of GUCA1A

• Calcium-sensitive regions of GCAP1 as observed by chemical modifications, fluorescence, and EPR spectroscopies [5].
• Successive inactivation of each of the functional EF loops by site-directed mutagenesis showed that only EF3 and EF4 contribute to a Ca2+-dependent inactivation of GCAP1 [16].
• Sequence analysis indicates that GCIP and GCAP1 and GCAP2 have diverged substantially, but conserved domains present in all vertebrate GCAP are present in GCIP [14].
• A mutation in GCAP1 segregating with autosomal dominant cone degeneration was further evaluated biochemically by employing recombinant proteins, immunoblotting, Ca2+-dependent stimulation of GC, fluorescence emission spectra, and limited proteolysis in the absence and presence of Ca2+ [2].
• Native and all three mutant forms of GCAP1 had similar affinities for Ca2+ as demonstrated by gel filtration and the changes in tryptophan fluorescence [17].

References